The present invention relates in general to an ion detector, and more particularly to an ion detector for selective and enhanced detection of large mass to charge ratio ions.
Ions having large mass to charge ratio (m/z) (ions greater than approximately 12,000 daltons) typically may be generated via several different ionization techniques, including but not limited to: Plasma Desorption/Ionization (PDI), Matrix-assisted Laser Desorption/Ionization (MALDI), Surface-enhanced Laser Desorption/Ionization (SELDI), and Electrospray Ionization (ESI). The large m/z values for these ions are such that they are beyond the m/z dynamic range for most simple magnetic sector, electrostatic analyzer, magnetic sector hybrid, and quadrapole filter analyzers. Consequently, the analysis of these ions is typically performed using ion-trap, fourier transform ion cyclotron resonance, and time-of-flight (TOF) mass spectrometers. Because of their simplicity and economy, when compared to the other previously mentioned devices, TOF systems are most frequently used of analyzing such large ions.
In time-of-flight methods of mass spectrometry, charged (ionized) molecules are produced in a vacuum and accelerated by an electric field produced by an ion-optic assembly into a free-flight tube or drift time. The velocity to which the molecules may be accelerated is proportional to the square root of the accelerating potential, the square root of the charge of the molecule, and inversely proportional to the square root of the mass of the molecule. The charged molecules travel, i.e., "drift" down the TOF tube to a detector.
FIG. 1 generally illustrates a laser desorption ionization time-of-flight mass spectrometer. Briefly, the system comprises ion optics 20, which include a repeller 21, an extractor 22, and a ground plate 23. A mass filter 24 may be included. A detector 25 completes the system. A crystallized layer of sample/matrix mixture 30 is applied to the surface of a probe 19. The ion optics are then energized and a laser beam 31 is applied to sample mixture 30 to thereby release or desorb ions. Repeller 21 is held at a potential of, for example, 30 kV, extractor 22 is held at a potential of, for example, 15 kV, while groundplate 23 is held at ground potential. An electric field is set up due to the potential difference between repeller 21, extractor 22, and groundplate 23, and thereby accelerates desorbed ions through the ion optics. Among the desorbed ions are matrix molecules and analyte molecules. Since the analyte molecules are the molecules of interest, mass filter 24 may be utilized to filter out the matrix molecules. Mass filter 24 typically comprises an entry plate and exit plate (not shown) and deflector. Finally, the ions reach detector 25 and the time-of-flight in traveling to the detector is utilized to calculate a mass to charge ratio. Since laser beam 31 passes through a beam splitter 27 such that a portion of laser beam 31 activates a trigger photo diode 32, the time the process started is known.
A laser desorption/ionization time-of-flight mass spectrometer (LDIMS), as depicted in FIG. 1, could be used to perform MALDI or SELDI analysis.
For MALDI analysis, samples are prepared as solid-state co-crystals or thin films by mixing them with an energy absorbing compound or colloid (the matrix) in the liquid phase, and ultimately drying the solution to the solid state upon the surface of an inert probe. In SELDI analysis, the probe or sample presenting surface plays an active role in the ionization, purification, selection, characterization or modification of the applied sample. In some cases an energy absorbing molecule (EAM) is an integral component of the sample presenting surface. In other cases, an energy absorbing molecule is added after the SELDI surface has completed its required interaction with the sample. Regardless of EAM application strategy, the probe contents are allowed to dry to the solid state prior to introduction into the LDIMS.
The output of detector 25 is integrated at some duty cycle as a function of time with respect to the time of the irradiating laser pulse 31 as sensed by the trigger photo diode 32. The molecular weight of an ion is then determined using the time-of-flight expression: m/z=A (Tf-To).sup.2 where: M/Z is the ions determined mass to charge ratio, Tf is the total flight time of the ion, To is the time interval that exists between the triggering of the timing device and acceleration of resultant ions and A is a constant that accounts for ion total kinetic energy and total flight distance. The values for A and To are empirically determined by comparing the experimental Tf flight number of well characterized analytes with their respective m/z. The determination of A and To calibrates the instrument and allows for more accurate m/z assignment.
During MALDI and SELDI analysis, a significant population of ions may be generated as a direct consequence of the use of matrix or EAM, respectively. These ions are transmitted down to the detector's conversion surface along with those ions created from the analytes of interest. In ESI analysis, a large number of ions are created from the solvents which make up the carrier solution. As was the case for SELDI and MALDI, these ions are also transmitted down to the detector's conversion surface. In all of these ionization techniques, it is not uncommon for these "unwanted ions" to be a major component of the entire ion current, far exceeding the number of analyte ions that are of interest. Since the ion transmission time period for a single LDIMS scan is rarely greater than 500 microseconds, detector electrons consumed during the conversion/gain process are usually not replaced during this rapid duty cycle. The result is charge depletion and field collapse to a level that seriously compromises detector gain.
In order to avoid field collapse and attendant gain reduction, presently used devices provide ways by which unwanted ions are prevented from striking the ion detector or ways by which detector gain voltage is rapidly switched on after the last unwanted ions strike the conversion surface. The former is accomplished by employing the additional set of ion optic elements that function as a mass gate or mass filter. The latter is accomplished through the use of high speed switching devices such as field effect transistors. Both of these methods add complexity and cost to TOFMS instruments. Because the gain rise time of a detector conversion surface is often several microseconds, the rapid switching technique does not allow for steep cut-off ranges, creating the possibility of inadequate gain during the initial phase of its duty cycle.
Ion detection in TOF mass spectrometry is typically achieved with the use of electro-emissive detectors such as electron multipliers (EMP) or microchannel plates (MCP). Both of these devices function by converting primary incident charged particles into a cascade of secondary, tertiary, quaternary, etc. electrons. The probability of secondary electrons being generated by the impact of a single incident charged particle can be taken to be the ion-to-electron conversion efficiency of this charged particle (or more simply, the conversion efficiency). The total electron yield for cascading events when compared to the total number of incident charged particles is typically described as the detector gain. Because generally the overall response time of MCPs is far superior to that of EMPs, MCPs are the preferred electro-emissive detector for enhancing m/z resolving power. However, EMPs function well for detecting ion populations of disbursed kinetic energies, where rapid response time and broad frequency band width are not necessary.
The conversion efficiency of large ions is known to be two to three orders of magnitude less than that of smaller ions. To compensate for this effect, secondary ion generators (SIG) have been used. Such a secondary ion generator is disclosed in U.S. Pat. Nos. 5,382,793, and 5,594,243, the contents of which is incorporated herein by reference for all purposes. With such secondary ion generators, when a primary incident ion strikes the surface of a secondary ion generator held at ground potential, secondary ions are created via the fragmentation of primary incident ions as well as the sputtering of what was thought to be a significant population of secondary metal ions from the SIG surface. FIG. 2a depicts an MCP detector utilizing a discrete SIG. In this arrangement, the SIG is a low transmission grid that is generally composed of copper or some copper alloy. It was postulated that incident ions (M+H)+strike the SIG resulting in their fragmentation into a series of product ions and neutrals as well as the release of electrons and SIG structural ions (in this example, Cu+). SIG product ions are post accelerated to the MCP conversion surface through the use of moderately strong electrical fields (.about.-1 to -5 kV/cm). Since the m/z of the SIG product ions are typically far less than that for large primary incident ions, ion conversion efficiency is increased and sensitivity can be improved by two--three orders of magnitude.
Recent work has led to the discovery that the majority of sputtered products from such secondary ion generators are actually emitted electrons and metal neutrals, and not a predominance of secondary metal ions as previously believed. Furthermore, it has been discovered that a significant population of these sputtered products are emitted in retrograde fashion with respect to the original direction of incident ion trajectory. FIG. 2c depicts this process.
It has also been discovered that biasing the SIG to some negative potential, such as -50 to -3,000 volts, improved the collision probability of primary ions by suppressing any strong "field punch," penetration of an electric field from one region into another, created by the underlying MCP conversion surface held at negative potentials greater than -2 KV. Such field punch provides an accelerating field which preferentially directs incident ions away from the SIG grid wires and into the space between them thus defeating the purpose of the SIG.
It has also been demonstrated that biasing the SIG to some negative potential promotes the emission of electrons. The emission of sputtered neutral products are not effected during such biasing. Since the negatively biased SIG is typically mounted upon an MCP detector whose impact surface is held at some high negative potential exceeding that which is employed in the SIG biasing, both forward and back sputtered electrons are accelerated backwards in retrograde fashion. Consequently, these electrons are driven through the cloud of sputtered neutrals, thus ionizing them to sputtered metal ions through the mechanism of electron impact ionization. In this manner, ion-converted, back sputtered neutrals can now be accelerated by the field of the negatively biased SIG to pass through the SIG and strike the surface of the MCP thereby creating additional detection signals which enhances the sensitivity for high molecular weight ions.
In addition to ionizing sputtered neutrals, such retrograde electrons promote fragmentation of non incident and soon to be incident parent ions through the mechanism of electron impact. Since the m/z of these fragment ions are less than that for their large, primary ions, ion conversion efficiency is further increased.
Since far more sputtered metal-neutral products are formed than sputtered metal ions, and because 2 significant population of these products are released in retrograde or back sputtered fashion, and because emitted electrons can be used to fragment primary ions or convert sputtered neutral products to forms more amenable to detection, and further because field penetration through a ground potential SIG reduces primary ion impact, prior art SIG approaches, as depicted in FIG. 2a, do not make optimum use of this secondary ion generation process. Significant improvement in the detection of high molecular weight ions can thus be achieved by negatively biasing the surface of the SIG. Because a biased SIG is more successful in generating charged, detectable products, such a configuration will now be referred to as a secondary charged particle generator (SCPG).
SIG or SCPG fragmentation, ionization, and sputter products are generated at a plurality of times, masses, and energies, and thus, many of these products do not propel uniformly forward and therefore do not strike the MCP conversion surface at the same time as their non-incident, parent ion counterparts. Therefore, discrete SCPG or SIG devices introduce ion conversion time spread and can result in an attenuation of m/z resolution. Such ion conversion time spread can be tolerated if it is insignificant when compared to other existing time spreads created in the measuring process. The initial ion energy spread of large ions are beyond the energy focusing capability of present SELDI and MALDI TOF technology, and is the limiting factor in m/z resolution. Consequently, a discrete SCPG or SIG can be used to increase ion conversion efficiency and detection sensitivity without significant reduction of m/z resolving power. However, for smaller ions, significant reduction of m/z resolving power has been demonstrated when using a discrete SCPG or SIG in these applications.
It has been demonstrated that the placement of an additional grid electrode (a differential acceleration grid, DAG) between the SCPG and MCP can be used to mitigate the time of flight disparity between SCPG generated products and non-incident parent ions thus improving mass resolving power. Such an arrangement is depicted in FIG. 2c. SCPG created sputter products are generally much lower in MW than their incident ion or fragmentation ion counterparts. Consequently, acceleration produced within the field that exists between the SCPG and MCP often propels sputtered ion products past these other ions. The result can range from a front end distorted detection signal to the resolution of early arriving ion populations, depending on the mass of the incident ion. Ions with MW less than 50 kDa can typically produce two or more measurable signals while heavier ions tend to have a single, front end distorted signal.
Such distortion of resolution could be avoided by placing a low acceleration potential between the SCPG and the MCP, however doing so will greatly reduce the final energy of sputtered and fragmented SCPG products, thus reducing their electron conversion efficiencies at the detector surface. Additionally, the use of high strength post acceleration fields have also demonstrated improvements in non-incident parent ion detection conversion efficiency, further augmenting sensitivity for large mw ions. Thus, it is advantageous to have strong acceleration fields between the SCPG and MCP surface.
A preferred method to eliminate this problem involves the use of a DAG. An electrical potential is placed upon the DAG which establishes a field between the SCPG and the DAG which is significantly lower than that which would normally exist between the SCPG and an MCP. In this manner sputtered product ions are not greatly accelerated. Because the initial energies of these sputtered product ions are low (measured to be less than 20 eV), they move slowly through this region. Non incident parent ions and incident ions without significant energy loss, continue to move at high velocities through this region, passing the sputtered product ions. Once sputtered product ions pass the DAG they are then accelerated by a strong field existing between the DAG and MCP surfaces. The DAG potential is selected such that further acceleration of sputtered ion and parent ion populations occurs in a manner so that sputtered product ions "catch up" with the parent ion population at the point of impact upon the MCP surface. In this manner, time spread is minimized and resolution is improved.
Because the degree of differential acceleration required to time compensate parent and sputtered product ions is mass dependent, the potential of the DAG must vary as the mass of the -incident ion varies. This can be achieved by the use of distinct DC DAG potentials so that scans are segmentally performed at different target masses. However, this technique is somewhat cumbersome. A preferred solution is one in which the DAG is held at some constant DC potential and is capacitively coupled to an AC signal whose amplitude is time dependent. The time-dependent amplitude change of this AC signal is synchronized with the time of parent ion arrival at the SCPG, so that the appropriate DAG potential is present during a given mw analysis time.